Innate immunity is an ancient defense response that evolved with the earliest metazoan creatures, and is the first line of defense against microbial infection. These responses rely on the recognition of microbes by germline-encoded receptors, and drive the production of numerous chemical, biological, and cellular defense responses. In the face of constant microbial assault, innate immunity is essential for the survival of nearly all multicellular organisms. On the other hand, over-exuberant or inappropriate innate immune responses are the underlying cause of morbidity and mortality associated with many infectious and autoimmune diseases. The endocrine system, through steroids as well as sex hormones and vitamin D, has profound pro- or anti- inflammatory effects on the innate immune response. This crosstalk between the innate immune and endocrine systems is found throughout the animal kingdom, and likely evolved with some of the earliest animals. This proposal uses the fruit fly Drosophila melanogaster as a model for the study of these interactions. Flies offer many advantages for these studies, including experimental tractability with arguably the most robust genetic system for in vivo studies, extensive knowledge of steroid hormone regulatory networks, and an innate immune system without the complexity of the adaptive immune response. In addition, the Drosophila immune response is an excellent model for vector insect species, and discoveries made in flies are being translated into new approaches to control vector-borne diseases. Furthermore, many aspects of the innate immune responses are highly conserved with mammals, and discoveries made in flies can be translated into paradigm shifting findings in mammals. Particularly relevant for this proposal are the conserved NF-?B signaling pathway, which drives the immediate response to infection, and the modulation of these signaling pathways by steroid hormones. Significantly, steroid signaling is highly conserved between flies and mammals. A thorough mechanistic analysis of how the innate immune response is regulated by steroid hormones in the Drosophila model system will provide a deeper understanding of these ancient regulatory interactions, and are likely to identify new avenues for manipulating these interactions in vector insects and/or mammals. Preliminary data demonstrate that the insect steroid hormone 20-hydroxyecdysone (ecdysone) has a profound enhancing effect on NF-?B dependent innate immune responses. Ecdysone appears to modulate the Drosophila innate response by at least two mechanisms, by controlling expression of a key innate immune receptor and by regulating induction of specific target genes. These results suggest that this steroid hormone functions to sculpt immune responses during development as well as to prime more efficient responses during times of stress. Aims 1 & 2 are designed to elucidate the molecular mechanisms underlying this hormonal control of immunity as well as probe its underlying function(s). In Aim 3, we will test a novel alternative hypothesis that steroid-regulated immune signaling and antimicrobial peptides (AMPs) production facilitate developmental cell death. PUBLIC HEALTH RELEVANCE: Innate immunity plays a critical role in nearly all infectious and autoimmune diseases, and is very similar in nearly all animals. Steroid hormone signaling is also very similar in throughout the animal kingdom. Thus, the innate immune system of the fruit fly Drosophila melanogaster and how it is modulated by steroid hormones will be investigated. This research may have a direct and profound impact on continuing efforts to modulate the immune response in mosquitoes, in order to reduce the transmission of vector-borne diseases, such as malaria or West Nile Virus. In addition, the discoveries from this research may have direct relevance to similar innate immune and hormone signaling pathways in humans.